Synthesis and Evaluation of Novel Triaryl Derivatives with Readthrough-Inducing Activity

Abstract

The readthrough mechanism, which skips the premature termination codon and restores the biosynthesis of the defective enzyme, is an emerging therapeutic tactic for nonsense mutation-related diseases, such as Hurler syndrome, a type of mucopolysaccharidosis. In the present study, novel triaryl derivatives were synthesized and their readthrough-inducing activities were evaluated by a luciferase reporter assay with a partial α-L-iduronidase (IDUA) DNA sequence containing the Q70X nonsense mutation found in Hurler syndrome and by measuring the enzyme activity of IDUA knockout cells transfected with the mutant IDUA gene. KY-516, a representative compound in which the meta position carboxyl group of the left ring of the clinically used ataluren was converted to the para position sulfamoylamino group, the central ring to triazole, and the right ring to cyanobenzene, exhibited the most potent readthrough-inducing activity in the Q70X/luciferase reporter assay. In Q70X mutant IDUA transgenic cells, KY-516 significantly increased enzyme activity at 0.1 µM. After the oral administration of KY-516 (10 mg/kg), the highest plasma concentration of KY-516 was above 5 µM in rats. These results indicate that KY-516, a novel triaryl derivative, exhibits potent readthrough-inducing activity and has potential as a therapeutic agent for Hurler syndrome.

Discovery of Novel Chromenopyridine Derivatives as Readthrough-Inducing Drugs

“…KY-516 was effective against the Q70X nonsense mutation, but less effective against the W402X nonsense mutation. 11) In addition, KY-516 was ineffective even at 30 µM in the Hurler patient-derived fibroblasts carrying the W402X mutation used in this study. Therefore, we investigated the sequence specificity of the readthrough-inducing activity of KY-640 using the previously reported luciferase assay system.…”

“…Therefore, we investigated the sequence specificity of the readthrough-inducing activity of KY-640 using the previously reported luciferase assay system. 11) The results obtained showed that the readthroughinducing activity of KY-640 was stronger against the W402X mutation than the Q70X mutation (data not shown), in contrast to that of KY-516. The reason for the difference in sequence specificities between KY-516 and KY-640 remains unknown; however, each of the previously reported readthrough-inducing compounds was previously shown to have different sequence specificities, which may also be affected by the peripheral sequence of a nonsense mutation.…”

“…IDUA enzyme activity restored by readthrough was evaluated using a fluorescent substrate as previously described. 11) The oral absorption of the synthesized compounds was evaluated by measuring the plasma concentrations of the compounds in male C57BL/6J mice after their oral administration at 30 mg/kg. All animal experiments in the present study were conducted according to the guidelines for animal experiments of our company and the guidelines for animal experimentation approved by the Japanese Association of Laboratory Animal Science.…”

“…We previously identified a compound (KY-516) with more potent readthrough-inducing activity than ataluren by structural optimization based on ataluren. 11) In the present study, we synthesized various derivatives based on the chromenopyridine skeleton, which we discovered in our skeletal discovery research, and evaluated their readthrough-inducing activities. Structure-activity relationships were discussed and a representative compound 30 with a basic fused ring structure was named KY-640 and selected for further evaluations.…”